(Phytophthora Spp.) in Major Citrus Growing Areas of Ethiopia

l of Hortic na ul r tu u r o e J Mekonen, et al., J Horticulture 2015, 2:4 Journal of Horticulture DOI: 10.4172/2376-0354.1000154 ISSN: 2376-0354

Research Article Open Access Assessing and Measuring of Citrus gummosis (Phytophthora spp.) in Major Citrus Growing Areas of Ethiopia Mekonen M*1, Ayalew A2, Weldetsadik K2 and Seid A2 1Ethiopian Institute of Agricultural Research (EIAR), Melkassa Agricultural Research Center, Adama, Ethiopia 2Haramaya University, P.O. Box 138, Dire Dawa, Ethiopia *Corresponding author: Mekanon Moges, Ethiopian Institute of Agricultural Research (EIAR), Melkassa Agricultural Research Center, Adama, Ethiopia, Tel: 251-222-250228, 251-913-386389; E-mail: [email protected], [email protected] Rec date: April 16, 2015; Acc date: July 17, 2015; Pub date: July 20, 2015 Copyright: © 2014 Mekanon M, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Abstract

Citrus production in Ethiopia is threatened by a number of biotic and abiotic factors. Among these citrus gummosis is one of the most important biotic constraints in the country. This study was conducted with the objective to measure and estimate the incidence and distribution of citrus gummosis (Phytophthora spp.) in major citrus growing areas of Ethiopia. Surveys were conducted in ten major citrus growing areas of Ethiopia. In each orchard, 30 trees were selected and totally 300 trees were assessed for gummosis incidence. These trees were sampled by making two diagonal transects across the field in the form of an “X”. A tree decline was scored on a 0-4 scale where 0=free of any decline symptom, 4=complete drying of the plant. A tree was defined and recorded as cankered when it had any of the following symptoms: discoloration of the bark surface, discoloration of the underlying tissues, dieback, dried the whole part of the plant and exudation of gum from infected tissues. Results showed that growth and fruit production are greatly reduced on trees infected by this disease. Cracked lesions that exude sap are found on infected scions, which become gradually girdled and killed. This symptom was prevailed in 90% of the surveyed citrus orchards. Gummosis was more frequent on scions (trunk and branch) which had 66.96% infection than rootstocks (33.02%). generally the issue needs more attention in the field management including harvesting method (avoid climbing on the tree rather through ladder), don’t touch the tree with the equipments and hands with mud probabily have an inoculums; when irrigating and weeding and finally use of fungicides when pruning of the diseased branch.

Keywords: Citrus; Assessment; Dieback; Gummosis; Phytophthora; Hence, there is a need to undertake coordinated and multi-phased Survey; Ethiopia research investigations on possible causes of citrus decline and find a sustainable, eco-friendly and easily commercialized technology that Introduction fits for our subsistence agriculture system. Determining the actual importance of a possible citrus decline factors viz gummosis Citrus (Citrus sinesis L.) is one of the most important fruit crops (Phytophthora spp. ) with the aim to develop eco-friendly and known by humans since ancient times and is a good source of vitamin sustainable measures for their management is an integral part of this C with high antioxidant potential [1]. The origin of citrus is believed to coordinated research effort. Foot rot and gummosis occur when be south eastern Asia including eastern Arabia to the Philippines and Phytophthora propagules are splashed onto the trunk near ground from the Himalayas south to Indonesia or Australia [2]. Citrus level, infect wounds or growth cracks and produce lesions which constitutes a major group of fruits comprising of mandarins, oranges, extend down to the bud union and cause a twig dieback and if severe, lemon, pummelo, grape fruits, tangelo, trifoliate orange, citron, and trees may eventually die [7]. Few studies have suggested a possible citranges [3]. Citrus is among the most important fruit crops in mechanism of gummosis occurrence on citrus tree and as a cause of Ethiopia. Its cultivation started in Upper Awash valley and Melkassa for citrus decline elsewhere in the world much work was not done in areas in central Ethiopia [4]. It is being produced mainly in Dire Dawa Ethiopia thus far on gummosis. Consequently this study was areas, lower and middle Awash and Melkassa areas in southeast region conducted with the objective to measure and estimate the incidence of Ethiopia. Ethiopia's agro-climatic conditions are highly suitable for and distribution of citrus gummosis (Phytophthora spp.) from major production of high quality citrus fruits. Five phenological regions were citrus growing areas of Ethiopia. identified based on the blooming season and climate as potential production centers capable of supplying citrus throughout the year [5]. Materials and Methods It occupied 7290 hectares of land in 1985 in the country, but the area has come down to 5,380 hectares (2200 oranges, 1750 mandarin and 1100 hectares limes and lemon) with a production of 33,500 metric Description of the study area tons only [6]. The decline in production land through time was A survey was conducted between October and November 2011 to attributed to a number of production limiting biotic and abiotic determine the incidence and distribution of gummosis (Phytophthora factors. spp.) in major citrus growing areas of Ethiopia. In all orchards, the root stock was sour orange and the scion was the sweet orange cultivar Valencia in all the surveyed orchards. Disease survey was conducted in

J Horticulture Volume 2 • Issue 4 • 1000154 ISSN:2376-0354 Horticulture, an open access journal Citation: Mekonen M, Ayalew A, Weldetsadik K, Seid A (2015) Assessing and Measuring of Citrus gummosis (Phytophthora spp.) in Major Citrus Growing Areas of Ethiopia. J Horticulture 2: 154. doi:10.4172/2376-0354.1000154

Page 2 of 4 ten established citrus orchards in Upper awash, Errer-Gota, and Koka sampling orchards. More than 70% of surveyed orchards the dieback areas. The sites were located at 9°54’ N, 37°43’ E and 1175 masl (upper symptoms of the tree were observed along with the gumming from awash), 10°54’N, 37°75’E and 1174 masl (Erer Gota) and 9°28’N, main branch (Figure 1). Branches with this symptom resulted in die 37°50’E and 1611masl (Koka) areas. The average minimum and back of a tree that could not keep their fruits and leaves alive for the maximum annual temperature were 15.3°C and 32.6°C at Upper next season. Alvarez et al. (2007) the disease Phytophthora gummosis Awash, 17°C and 29°C Erer-Gota. causes necrosis of the inner bark and the cambium of the trunk and Phytophthora root rot, foot rot also may cause tree decline in severe Disease assessment cases tree wilt and death. In each orchard, 30 trees were selected and a gummosis incidence was assessed. These trees were sampled by making two diagonal transects across the field in the form of an “X” (15 plants along each diagonal). Gummosis incidence was determined as the proportion of plants showing gummosis symptoms, expressed as a percentage of the total number of plants assessed [8]. A tree was defined and recorded as cankered when it had any of the following symptoms: discoloration of the bark surface, discoloration of the underlying tissues, dieback, dried the whole part of the plant and exudation of gum from infected tissues. Tree decline was scored on a 0-4 scale where 0=free of any decline symptom, 4= complete drying of the plant (Table 1). Disease severity Figure 1: (A) Gummosis on the main branch of sweet orange (citrus index (DSI) was then calculated as: sinensis) tree and (B) tree dieback along the gummosis symptom on the main branch at Hurso 2011(Photo courtesy of Moges Mekonen). Disease index for severity DSI Disease scale Descriptionsum of all disease ratings of disease status 0Tree with no symptom =Total number of assessed plants x maximum rating value×100 Erer and Hurso the disease incidence reaches up to 100% while at associated with gummosis1Decline symptom associated with Tibilla 0.0% were recorded at the time of survey (Table 2). In the root gummosis up to 25% of the branch affected 2widespread decline of the stock the highest gummosis frequency were recorded at Erer (70%) branch associated with gummosis up to 25-50% of the branch affected followed by Hurso (46.67%). However, at the trunk as well as the 3Decline and death of the branch associated with gummosis up to branch the highest frequency were recorded at Hurso with a 90% and 50-75% of the branch affected4Decline of 75-100% tree, including dead 63.33% followed by Erer 83.33%, and 53.33% frequency respectively. tree. No Locations Orchard in Sample Incidence Tree decline Isolation of Phytophthora spp. from bark size (%) a Samples with underlying tissue were excised using a sharp knife 1 Dire Dawa Tony farm 30 60 1.6 which was surface disinfected through wiping with ethyl alcohol before 2 Hurso Hurso 30 100 2.875 and after each use. The disease samples were taken from the active lesion near to the crown and branch showing the symptom of the 3 Errer Gota Erer 30 100 3.642 disease. A total of 30 samples, obtained from 10 surveyed orchards, Gota 30 26.7 2.375 were processed. Pieces of 3-5 mm-long tissue were transferred using sterile forceps to a Petri dish containing modified V-8 agar medium Fetule 30 86.7 2.625 and incubated at 23°C in the dark and examined within 3–5 days. The modified V-8 agar consisted of eight vegetable agar (V-8 agar) 4 Upper Nura Era 30 13.3 2.092 Awash composed of 2 g CaCO3, 200 mL V8 juice and 15 g agar in 800 mL distilled water amended with 20 mg Pimaricin, 150 mg Ampicilin, 75 Merti 30 16.7 1.53 mg Vancomycin, 50 mg Dichloran, which were dissolved in 10 ml distilled sterile water and added to 1 liter of V-8 agar [10]. Pure Tibilla 30 0 0.917 cultures of Phytophthora spp. were obtained by transferring hyphal 5 Melkassa Melkassa 30 33.3 1.167 tips onto potato dextrose agar (PDA) which was used for colony pattern description, while V8 juice agar was used for morphological 6 Koka Koka area 30 13.3 2.525 description. Mean 30 45% 2.14 Results and Discussion Table 2: Incidence of citrus gummosis in ten major citrus growing orchards of Ethiopia, 2012. atree decline on a 0-4 scale where 0= no Distribution of citrus gummosis (phytophthora spp.) symptom and 4=decline of the tree 75-100%, including dead tree [9]. The survey results have shown that the disease was prevalent in all the surveyed orchards with different magnitude of infection. The Infections of tree parts gumming and cracking symptoms were observed on root stock, trunks Gummosis was more frequent on scions (trunk and branch) 66.96% and branches which produced a light yellow and brown gum across the than rootstocks 33.02% on the surveyed orchards. And from the scions

Page 3 of 4 cankers/gummosis were frequently recorded from the trunk 44.81% mean when altitude increases incidence on root stock, branch and than the branch 22·17% (Table 3). Alvarez et al. [7] reported that the trunk as well as tree decline also decreases. The citrus gummosis was citrus Cankers were more frequent on scions than rootstocks 92·4 and observed in 79% of the 90 orchards visited with high prevalence being 7·6% of respectively. The disease mainly affected mature citrus trees; observed in low altitude areas [16]. And when trunk incidence Diseased trees showed cankers and gum exudations mainly on above- increases branch incidence also increases which indicates that citrus union parts, especially on the major limbs, whereas rootstocks gummosis can affect branches if appropriate control measures aren’t generally remained healthy. timely designed.

Root stocka Location Trunkb (%) Branchc (%) (%) Disease severity index

Tony farm 13 14 - The overall result of disease severity index showed that Erer and Hurso orchards were highly affected by the disease. The lower disease Hurso 14 27 19 severities were recorded at Tibilla (22.917%) followed by Melkassa (29.17%) and Merti (38.33%) and the highest disease severity recorded Erer 21 25 16 at Erer (91.04%) followed by Hurso (71.89%), Fetule (65.625%) and Gota 1 7 1 Koka (63.125%) (Figure 2). Hebb and Sonoda [16] reported that in Florida a survey of 18 grape fruit groves showed that 3% to 74% of the Fetule 13 12 7 trees were affected by citrus gummosis. Similarly Jagtap et al. [15] also

Nura Era - 3 1 reported that in Marathwada region of Maharashtra state, all 103 sweet orange orchards surveyed had shown an average disease incidence of Merti 2 1 2 38.83%.

Tibilla - - -

Melkassa 3 6 -

Koka area 3 - 1

Total 70 95 47

Percentaged 33.02 44.81 22.17

Table 3: Occurrence of Phytophthora-cankers on root stock, trunk and branch of citrus trees in ten major citrus growing orchards of Ethiopia, 2012. aRootstock from ground to union line; bTrunk refers to main stem/branch; c all branches and sub-branches from the trunk and percentages of 212 infected trees of 10 surveyed orchards. Figure 2: The disease severity index from ten major citrus growing orchards of Ethiopia, 2012. The dominant status of P. citrophthora compared to other Phytophthora species on citrus is favored by environmental conditions [11,12]. The outbreak of this disease could be the variation in In the soil analysis result all citrus orchards show clay texture except environmental factors and changes in citrus cultural practices. Melkassa which is loam relatively with low PH (7.52). This clay nature Inoculums raised from the soil and move to the trunk and branches of the soil favors the pathogen by keeping the soil moisture for the long possibly due to poor cultural practice conducted at the orchards during period of time and breaking of spore dormancy to be infective. harvesting, irrigating and pruning. Likewise, the particular and most Leonard and Nathan [17] reports about the spore survival mechanism importantly this pathogen were spread during harvesting when were that the fungus survives in the soil as a thick-walled spore capable climbing to the tree. Since harvesting is conducted without ladder of withstanding extremes in both moisture and temperature. Clay when the harvester tries to climb to harvest he/she holds the nature of the citrus orchard and optimum temperature for the inoculums with the soil on their feet’s and hands. During irrigation, development of the spores of the pathogen as well as weak weeding, pruning etc. actions fever the pathogen to reach on the management of the orchards made the disease highly sever in the area. branch of the tree. In addition; probably some insects and animals have had contribute for the spread of the pathogen to the upper branch. The Incubation and isolation disease usually attacks the plant when soil comes into contact with the It was consistently isolated from diseased barks on modified scion, or when a tree is planted in a basin which may be flooded during selective medium PAVD (Figure 3). The Phytophthora spp. grown on irrigation, allowing the fungus to reach the scion [13-15]. V-8 medium and PDA produced sporangia abundantly in soil extracts. Morphological aspects of isolates were used in accordance with the Correlation between tree decline and altitude descriptions made by Drenth and Sendall [18] (Figure 3). Isolates from The analysis result showed that branch incidence was highly and the bark tested for growth on the selective medium at 23oc of positively correlated with trunk incidence (r=0.889), root stock temperature shows a white cottony colony after 72 hrs. But after seven (r=0.776). Root stock incidence was strongly and negatively correlated days of incubation different types of spores were become visible on the with altitude (r=-0.626). Altitude was also negatively correlated with culture under microscope including chlamydospores, sporangia and tree decline (r=-0.319) with weak association. These relations clearly hyphal swellings. Isolations from the bark of citrus tree revealed the presence of the fungus Phytophthora spp, the cause of citrus

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